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A Robust Single-Particle Cryo-Electron Microscopy (cryo-EM) Processing Workflow with cryoSPARC, RELION, and Scipion
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A Robust Single-Particle Cryo-Electron Microscopy (cryo-EM) Processing Workflow with cryoSPARC, RELION, and Scipion

A Robust Single-Particle Cryo-Electron Microscopy (cryo-EM) Processing Workflow with cryoSPARC, RELION, and Scipion

DOI:
10.3791/63387-v

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13:43 min

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January 31, 2022

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,
1Institute for Quantitative Biomedicine,Rutgers University, 2Department of Biochemistry & Microbiology,Rutgers University

Chapters

  • 00:04Introduction
  • 00:56Creating a New CryoSPARC v3 Project and Importing Data
  • 01:34CryoSPARC v3: Movie Alignment and Contrast Transfer Function (CTF) Estimation
  • 02:52CryoSPARC v3: Manual and Template-Based Particle Picking
  • 04:21CryoSPARC v3: 2D Classification
  • 04:52CryoSPARC v3: Ab-initio Reconstruction and Homogeneous Refinement
  • 05:31Exporting Particle Coordinates from CryoSPARC v3 and Importing Them to RELION-3 Using PyEM
  • 05:56RELION-3: Particle Extraction and 2D Classification, 3D Refinement, Mask Creation, and Post-Processing
  • 07:41RELION-3: Polishing Training, Particle Polishing, CTF, and Per-Particle Refinements
  • 09:15Transferring RELION-3 Particle Coordinates and 3D Map to Scipion 3
  • 10:05Scipion 3: High-Resolution Refinement
  • 11:21Results: Using cryoSPARC v3, RELION-3, and Scipion 3 to Obtain a High-Resolution 3D Reconstruction of the Adeno-Associated Virus
  • 12:52Conclusion

Summary

자동 번역

자동 번역

This article describes how to effectively utilize three cryo-EM processing platforms, i.e., cryoSPARC v3, RELION-3, and Scipion 3, to create a single and robust workflow applicable to a variety of single-particle data sets for high-resolution structure determination.

Tags

Keywords: Cryo-EMSingle-particle AnalysisCryoSPARCRELIONScipionAdeno-associated VirusGene TherapyPatch Motion CorrectionCTF EstimationMicrograph CurationManual Particle PickingTemplate GenerationParticle Extraction

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